bims-tuinly Biomed News
on Tumor-infiltrating lymphocytes therapy
Issue of 2026–09–06
nineteen papers selected by
Pierpaolo Ginefra, Ludwig Institute for Cancer Research



  1. J Vis Exp. 2026 Sep 03.
      Understanding the functional capacity of tumor-infiltrating lymphocytes (TILs) to recognize and eliminate autologous tumor cells is central to advancing personalized immunotherapy. The goal of this method is to provide an image-based, live-cell imaging protocol that measures TIL-mediated, caspase-3-dependent apoptotic killing against patient-derived tumor organoids (PDTOs) in real time. This method integrates established procedures for isolation and expansion of PDTOs and TILs with a standardized three-dimensional co-culture system and automated fluorescence-based apoptosis detection. Tumor organoids are plated in imaging-compatible 96-well plates and labeled with a red tumor marker, while expanded TILs are added at defined effector-to-target ratios in the presence of a caspase-3 activated green fluorescent substrate. Co-cultures are imaged every 4 h using a live-cell analysis system to capture phase-contrast and dual-fluorescence channels. Quantitative image analysis identifies red-positive tumor structures and calculates the proportion of red/green double-positive apoptotic tumor objects over time. Appropriate technical and biological replicates are incorporated, along with baseline, spontaneous apoptosis, negative and positive killing controls to ensure assay rigor. By preserving tumor heterogeneity within the PDTOs' three-dimensional architecture while enabling longitudinal quantification, this protocol provides a physiologically relevant system for functionally profiling patient-specific tumor-TIL interactions and investigating immunomodulatory agents that augment anti-tumor immunity.
    DOI:  https://doi.org/10.3791/71688
  2. Immune Netw. 2026 Aug;26(4): e31
      Cancer immunotherapy has reshaped oncology, largely through immune checkpoint inhibitors that release the brakes on tumor-reactive T cells. Yet the benefit remains uneven, and that unevenness traces back to a few basic biological limits. Checkpoint blockade amplifies immunity that is already present; it does not create tumor specificity de novo. Poor Ag quality, defective Ag presentation, a suppressive microenvironment, and epigenetically fixed T-cell exhaustion together set a ceiling on what checkpoint release can achieve. Next-generation strategies try to move past these limits by reorganizing immunotherapy around the functional layers of the immune response. Cancer vaccines define tumor-specific neoantigens and expand the responses against them. Ab-based approaches tune inhibitory signaling, draw immune cells toward the tumor, and trigger immunogenic cell death. Cellular therapies-chimeric Ag receptor T cell, TCR-engineered T cells, and tumor-infiltrating lymphocytes (TILs)-boost effector potency, with TIL therapy notable for preserving tumor-reactive repertoires shaped in vivo. Rather than rivals, these modalities are best seen as complementary layers-Ag definition, immune priming, effector optimization, and microenvironmental conditioning-to be combined in a programmable way. As genomic profiling, immunopeptidomics, and high-dimensional immune monitoring mature, the field is shifting from checkpoint-centered release toward precision immunoengineering, in which tumor-specific immunity is deliberately designed, aligned, and sustained.
    Keywords:  Cancer vaccines; Cell therapy; Immune checkpoint inhibitors; Immunotherapy; Neoantigens
    DOI:  https://doi.org/10.4110/in.2026.26.e31
  3. Crit Rev Oncol Hematol. 2026 Sep 01. pii: S1040-8428(26)00464-6. [Epub ahead of print]227 105577
      Relapsed or refractory neuroblastoma (R/R NB) is one of the most challenging pediatric cancers, with long-term survival rates below 20%. Both standard chemotherapy and single-target immune cell therapies are associated with significant limitations. Chemotherapy is unable to clear minimal residual disease in the majority of cases, and engineered cell therapies lose efficacy when tumors drop the target antigen because of intratumoral heterogeneity. Thus, tumor-infiltrating lymphocyte (TIL) therapy offers an alternative approach. TILs are polyclonal and have previously engaged with the tumor, so they can recognize a broad spectrum of patient-specific antigens and are less likely to be compromised by the loss of a single antigen. Nevertheless, early efforts to evaluate TIL therapy in R/R NB have encountered important challenges. The primary obstacles include unsuccessful TIL manufacturing, immunosuppressive tumor microenvironment (TME), and defective antigen presentation due to MYCN-driven loss of MHC class I and impaired interferon signaling. These challenges necessitate novel approaches, such as antigen enrichment or MHC-independent engineering. This review summarizes the results of preclinical research over three decades, categorizing studies into three major periods with distinctive determinants of TIL therapy efficacy in NB. Key clinical studies are also summarized to inform the development, safety, and feasibility of TIL therapy in pediatric NB patients. Finally, a translational roadmap is proposed that integrates γδ TIL enrichment, cell engineering, TME modification, and biomarker-driven patient selection to guide early-phase clinical trials in R/R NB.
    Keywords:  Adoptive cell therapy; Cell-intrinsic T cell engineering; Immunologically cold tumor microenvironment; Neuroblastoma; Pediatric solid tumors; Tumor-infiltrating lymphocytes; γδ T cells
    DOI:  https://doi.org/10.1016/j.critrevonc.2026.105577
  4. ASM Case Rep. 2026 Sep;pii: e00070-26. [Epub ahead of print]2(5):
       Background: Hemophagocytic lymphohistiocytosis (HLH) is a severe hyperinflammatory syndrome triggered by infections, malignancies, or autoimmune disorders. Tumor-infiltrating lymphocyte (TIL) therapy requires lymphodepleting chemotherapy and interleukin-2, producing profound immunosuppression. While HLH-like syndromes have been characterized after chimeric antigen receptor T-cell therapy, HLH has not been reported in the setting of TIL therapy. Toxoplasma gondii is a recognized but uncommon parasitic trigger of secondary HLH.
    Case Summary: A 68-year-old male with metastatic melanoma undergoing TIL therapy presented with altered mental status. He had chronic lymphopenia and reported gastrointestinal intolerance to trimethoprim-sulfamethoxazole, documented as a sulfonamide allergy without formal testing, limiting prophylaxis to inhaled pentamidine, which has no anti-Toxoplasma activity. On admission, he was febrile (39.3°C) with pancytopenia, hypofibrinogenemia (44 mg/dL [normal 200-400]), hyperferritinemia (>30,000 ng/mL [normal 30-300]), hypertriglyceridemia (410 mg/dL [normal 150]), and elevated soluble CD25 (6,979.4 pg/mL [normal 175.3-858.2]); HScore was 283 (>99% probability of HLH). Bone marrow biopsy revealed bradyzoites, tachyzoites, pseudocysts, and hemophagocytosis. Disseminated toxoplasmosis was confirmed by positive serum Toxoplasma IgG, negative IgM, and cerebrospinal fluid Toxoplasma PCR of 13,700 copies/mL (reference: not detected), with brain MRI demonstrating targetoid enhancing lesions. Treatment with pyrimethamine, sulfadiazine, and leucovorin was initiated, but the patient continued to deteriorate and transitioned to comfort measures.
    Conclusion: This represents the first toxoplasmosis-associated HLH during TIL therapy. It underscores the importance of distinguishing drug intolerance from true allergy, ensuring Toxoplasma-active prophylaxis in profoundly immunosuppressed patients, and recognizing that HIV-derived CD4 thresholds may not apply to non-HIV immunocompromised populations.
    Keywords:  HLH; TIL therapy; immunocompromised; opportunistic infection; toxoplasmosis
    DOI:  https://doi.org/10.1128/asmcr.00070-26
  5. Anticancer Res. 2026 Sep;46(9): 5095-5118
       BACKGROUND/AIM: Esophageal squamous cell carcinoma (ESCC) exhibits substantial heterogeneity in its tumor immune microenvironment (TIME). We aimed to establish an integrated spatial immune phenotyping framework incorporating immune infiltration, spatial distribution, and suppressive-effector immune balance.
    PATIENTS AND METHODS: A total of 139 patients with surgically resected ESCC were retrospectively analyzed, including a discovery cohort (n=89) and an external validation cohort (n=50). Whole-slide histopathological evaluation and immunohistochemistry were used to assess spatial immune-cell infiltration. Tumors were classified according to total tumor-infiltrating lymphocyte (TIL) level, the intratumoral-to-stromal TIL ratio, and stromal suppressive-to-effector immune balance. TCGA-ESCA transcriptomic data were analyzed to characterize the identified phenotypes.
    RESULTS: Stromal TILs significantly exceeded intratumoral TILs (p<0.001). Increased intratumoral immune infiltration and enrichment of suppressive immune-cell populations were associated with aggressive clinicopathological features. Four immune phenotypes were identified: immune-cold, immune-excluded, CD8-effector infiltrated, and suppressive-infiltrated. The suppressive-infiltrated phenotype showed the poorest overall survival in both the discovery (p=0.010) and validation cohorts (p=0.008). In the pooled cohort, Type IV status remained independently associated with worse overall survival after adjustment for major clinicopathological factors [hazard ratio (HR)=2.64, 95% confidence interval (CI)=1.20-5.79, p=0.015]. Adding Type IV status improved prognostic performance (C-index: 0.650 to 0.673; likelihood-ratio test, p=0.018). Transcriptomic analyses demonstrated increased immune checkpoint activity, T-cell exhaustion, and stromal activation signatures in the suppressive-infiltrated phenotype (all p<0.05).
    CONCLUSION: Integrated assessment of immune infiltration, spatial distribution, and suppressive-effector balance identified clinically distinct immune phenotypes in ESCC. The suppressive-infiltrated phenotype represents a high-risk immune microenvironmental subgroup independently associated with poor survival.
    Keywords:  Esophageal squamous cell carcinoma; immune phenotyping; immunosuppression; prognosis; tumor microenvironment; tumor-infiltrating lymphocytes
    DOI:  https://doi.org/10.21873/anticanres.18357
  6. Breast Cancer Res Treat. 2026 Aug 31. pii: 3. [Epub ahead of print]219(2):
       BACKGROUND: Neoadjuvant chemotherapy, a cornerstone of early-stage triple-negative breast cancer (TNBC) treatment, has limited efficacy and is often accompanied by substantial toxicity. As a promising chemotherapy-free alternative, the combination of immune checkpoint inhibitors and anti-angiogenic agents provides the potential to enhance antitumor efficacy by remodeling the tumor microenvironment while reducing treatment-related adverse events (TRAEs). This study aimed to assess the clinical value and feasibility of neoadjuvant camrelizumab plus apatinib in patients with early-stage TNBC.
    METHODS: This study enrolled stage II-III TNBC patients with baseline tumor-infiltrating lymphocytes (TILs) > 10%. Participants received 8 cycles of neoadjuvant treatment, consisting of intravenous camrelizumab 200 mg every 21 days (or 3 mg/kg for patients weighing < 50 kg), combined with oral apatinib 250 mg daily. The primary endpoint was pathological complete response (pCR) rate; secondary endpoints included objective response rate (ORR), disease-free survival (DFS), overall survival, and safety.
    RESULTS: Between December 2022 and March 2024, 25 patients were screened, 14 were recruited and treated, 13 were evaluable for efficacy, and 11 subsequently underwent surgery. The results showed a pCR rate of 27.3% (3/11), an ORR of 46.2% (6/13), and a 24-month DFS rate of 90.9%. TRAEs were predominantly of grade 1-2, with grade ≥ 3 events occurring in 28.6% patients. Exploratory transcriptomic analysis identified differences in genes related to extracellular matrix remodeling and immune regulation.
    CONCLUSIONS: Camrelizumab plus apatinib demonstrated manageable toxicity and preliminary antitumor activity, supporting further investigation of this chemotherapy-free strategy in early-stage TNBC patients with enriched TILs.
    TRIAL REGISTRATION: NCT05556200 (Registration Date: September 27, 2022).
    Keywords:  Anti-angiogenic therapy; Immune checkpoint inhibitors; Neoadjuvant therapy; Triple-negative breast cancer
    DOI:  https://doi.org/10.1007/s10549-026-08066-5
  7. Exp Mol Med. 2026 Aug;58(8): 2590-2602
      T cell exhaustion arises during chronic antigen stimulation and represents a major barrier to effective anti-tumour immunity. Rather than a uniform dysfunctional state, exhaustion is increasingly understood as a structured differentiation landscape that progresses from stem-like progenitor exhausted T cells to terminally exhausted T cells. Stem-like progenitor exhausted T cells retain self-renewal capacity and partial effector function, whereas terminally exhausted T cells exhibit epigenetically fixed dysfunction and limited cytokine production. Within the tumour microenvironment, persistent antigen stimulation, together with metabolic stressors such as hypoxia and nutrient deprivation, accelerates this differentiation trajectory, thereby constraining protective immunity. In this review, we conceptualize T cell exhaustion as a dynamic continuum shaped by both differentiation states and microenvironmental niches. We outline an integrative framework to define exhaustion by combining antigen experience, cellular phenotype, functional capacity, epigenetic fixation and spatial context. Viewing immunotherapies through this multidimensional framework highlights the notion that durable therapeutic responses depend on preserving stem-like progenitor exhausted T cells within supportive niches and preventing their terminal differentiation. Understanding how these cellular states are maintained or disrupted within the tumour microenvironment will provide new opportunities for designing therapies that sustain protective T cell immunity in cancer.
    DOI:  https://doi.org/10.1038/s12276-026-01809-w
  8. Int J Biol Sci. 2026 ;22(13): 7053-7081
      CD8⁺ T-cell exhaustion is a distinct differentiation state driven by persistent antigen stimulation, and its establishment and maintenance are major barriers to effective cancer immunotherapy. Although the transcriptional and epigenetic landscapes of exhausted CD8⁺ T cells have been extensively characterized, it remains unclear how sustained external stimulation is integrated at the level of protein function to produce stable dysfunction and altered cell fate. Post-translational modifications constitute a key regulatory layer of protein function. They form a dynamic network that links persistent antigenic stimulation and tumor microenvironmental stress to cell fate, and therefore provide a critical entry point for understanding how exhaustion is initiated and maintained. In this review, we focus on how post-translational modifications convert persistent antigen stimulation and tumor microenvironmental stress into protein-level dysregulation and ultimately lock CD8⁺ T cells into an exhausted fate. We summarize how multiple classes of post-translational modifications drive exhaustion through effects on signal transduction, protein homeostasis, metabolic stress responses, and epigenetic reprogramming. We then discuss potential intervention strategies centered on critical regulatory nodes that may preserve the plasticity of precursor exhausted CD8⁺ T cells, restrain stabilization of the terminally exhausted state in CD8⁺ T cells, and optimize rational combination therapies. Finally, we outline the translational challenges and future directions of targeting post-translational modifications, and emphasize that identifying actionable modification nodes will be important for patient stratification and combination design in cancer immunotherapy.
    Keywords:  CD8⁺ T-cell exhaustion; cancer immunotherapy; immune checkpoint blockade; post-translational modifications; tumor microenvironment
    DOI:  https://doi.org/10.7150/ijbs.138638
  9. Med. 2026 Sep 02. pii: S2666-6340(26)00275-8. [Epub ahead of print] 101272
       BACKGROUND: Mucosal-associated invariant T (MAIT) cells are innate-like T lymphocytes that recognize riboflavin-derived metabolites presented by the monomorphic major histocompatibility complex (MHC)-class I-related molecule MR1. MAIT cells are highly enriched in the human liver and are increasingly implicated in liver cancer immunity, yet their antitumor potential and therapeutic applicability remain incompletely defined.
    METHODS: Here, we profiled MAIT cells derived from peripheral blood of healthy donors and patients with hepatocellular carcinoma, revealing disease-associated phenotypic alterations marked by immune checkpoint dysregulation. To address current therapeutic limitations, we developed a robust ex vivo expansion platform capable of generating MAIT cells with high yield, purity, and potent cytotoxic function. Using multiple human liver cancer subcutaneous and orthotopic xenograft mouse models, we demonstrate that adoptively transferred MAIT cells mediate effective tumor targeting and killing in vivo.
    FINDINGS: Mechanistically, MAIT cell antitumor activity is driven by both T cell receptor (TCR)-dependent recognition and natural killer receptor (NKR)-mediated cytotoxicity and is further enhanced by antigen stimulation; cytokine receptor signaling, particularly via interleukin (IL)-15; and immune checkpoint modulation involving programmed cell death protein 1 (PD-1) and T cell immunoglobulin and ITIM domain (TIGIT). In addition, MAIT cells demonstrated a favorable safety profile, with no evidence of liver toxicity observed in xenograft mouse models.
    CONCLUSIONS: Together, these findings establish MAIT cells as functional antitumor effectors in liver cancer and provide a mechanistic and translational framework for adoptive MAIT cell therapy enhanced by multimodal modulation of immune receptors and checkpoints.
    FUNDING: Major funding was provided by the California Institute for Regenerative Medicine (CIRM).
    Keywords:  5-OP-RU; MAIT; PD-1; TIGIT; cancer immunotherapy; cytokine receptor; hepatocellular carcinoma; immune checkpoint; liver cancer; mucosal-associated invariant T cell; pre-clinical research
    DOI:  https://doi.org/10.1016/j.medj.2026.101272
  10. Nat Chem Biol. 2026 Sep 01.
      Cell-surface sialoglycans overexpressed on cancer cells suppress activation of tumor-infiltrating immune cells (TIICs) and thus represent promising targets for cancer immunotherapy. However, the functional roles of sialylation in TIICs remain incompletely understood. Here we developed a strategy for single-cell correlative analysis of α2,3- and α2,6-linked sialoglycans and transcriptomes to determine the correlations of cell-surface sialylation with gene expression of the same cells in a linkage-specific manner by single-cell RNA sequencing. We found that α2,3- and α2,6-sialoglycans were distinctively regulated across specific TIIC subpopulations in murine tumors. Notably, downregulation of α2,6-sialoglycans was identified as a cell-surface marker for intratumoral tumor-specific antigen-reactive CD8+ T cells with high avidity. Mechanistically, high-avidity (but not low-avidity) activation of murine CD8+ T cells within the tumor suppressed β-galactoside α2,6-sialyltransferase 1 expression, which downregulates α2,6-sialylation and exposes galectin-1 ligands. Functionally, galectin-1 binding promoted T cell apoptosis and immune evasion in cancer. This work demonstrates the functional significance and therapeutic potential of specific sialoglycans on TIICs.
    DOI:  https://doi.org/10.1038/s41589-026-02299-7
  11. Biomark Res. 2026 Aug 11. pii: 115. [Epub ahead of print]14(1):
       BACKGROUND: Although chimeric antigen receptor T-cell (CAR-T) therapy has reshaped the treatment landscape for relapsed/refractory multiple myeloma (RRMM), durable remission remains challenging. T-cell dysfunction and exhaustion are key obstacles to long-term efficacy. Programmed death-ligand 1 (PD-L1)-positive exosomes may contribute to tumor-associated immunosuppression, but their clinical and functional relevance in multiple myeloma (MM) remains unclear.
    METHODS: A total of 89 patients with MM were enrolled, including 34 patients with RRMM treated with CAR-T therapy. PD-L1+ exosome levels in peripheral blood (PB) and bone marrow (BM) were quantified by flow cytometry using exosome-coupled beads, and soluble PD-L1 (sPD-L1) was measured by enzyme-linked immunosorbent assay. Associations with clinicopathological features, treatment response, and progression-free survival (PFS) were analyzed. Effects of RRMM patient sample-derived PD-L1+ exosomes on CD8+ T-cell activation, cytokine production, and exhaustion phenotypes were assessed using in vitro experiments. The therapeutic potential of GW4869 combined with anti-programmed cell death protein 1 (PD-1) antibody to enhance CAR-T efficacy was further evaluated using in vitro and in vivo models.
    RESULTS: PD-L1+ exosome levels in PB and BM were significantly elevated in newly diagnosed MM and RRMM compared with healthy controls, and were closely associated with adverse prognostic features, including high tumor burden, extramedullary lesions, and high-risk cytogenetic features, whereas plasma sPD-L1 showed no significant clinical correlations. In RRMM patients receiving CAR-T therapy, pretreatment BM and PB PD-L1+ exosome levels discriminated best response (complete response versus less than complete response), with BM showing better predictive performance (area under the receiver operating characteristic curve, 0.846; optimal cutoff, 65.0%). High pretreatment PD‑L1+ exosome levels were associated with low peak levels of CAR transgene, interferon-γ, and tumor necrosis factor-α, as well as shorter PFS. In addition, elevated BM PD‑L1+ exosome level remained an independent adverse factor for PFS in multivariable analysis. In vitro, RRMM patient sample-derived PD-L1+ exosomes inhibited CD8+ T cell activation and effector cytokine production, and promoted terminal exhaustion, which was not fully reversed by anti-PD-1 antibody alone. In preclinical models, GW4869 plus anti-PD-1 antibody enhanced CAR-T-mediated antitumor activity in both cell-based assays and the MM xenograft model.
    CONCLUSION: PD‑L1+ exosomes may serve as biomarkers of disease aggressiveness and CAR‑T therapy outcomes in MM. By impairing CD8+ T cell function and promoting terminal exhaustion, they may contribute to immune resistance. Combined exosome-release pathway and PD‑1 blockade warrants further investigation as a strategy to enhance CAR‑T efficacy.
    Keywords:  Chimeric antigen receptor T cell; Combination therapy; Multiple myeloma; PD-L1+ exosomes
    DOI:  https://doi.org/10.1186/s40364-026-00980-6
  12. Nat Med. 2026 Sep 01.
      Triple-negative breast cancer (TNBC) is an aggressive subtype with an activated tumor immune microenvironment. The multicenter, multinational, double-blinded NSABP B-59/GeparDouze trial evaluated the addition of atezolizumab (atezo) (773 patients randomized) or placebo (777 patients) to sequential taxane-carboplatin-anthracycline-based neoadjuvant chemotherapy in stage II-III TNBC. The addition of atezo did not significantly improve the primary endpoint of event-free survival (EFS) (HR, 0.80 (95% CI, 0.062-1.03); stratified log-rank P = 0.083, 4-year EFS rates difference 3.3%). The HR for overall survival was 0.86 (95% CI, 0.62-1.19), with a 4-year benefit of 0.7%. Prespecified subgroup analyses suggested heterogeneity in EFS, with benefit of atezo in patients presenting with clinical lymph node involvement (Pinteraction = 0.039). Treatment-emergent adverse events with grades ≥3 were reported in 75.3% (atezo) versus 73.4% (placebo), and immune-related adverse events were reported in 27.6% (atezo) versus 11.4% (placebo). A total of 196 (25.5%) patients discontinued atezo and 143 (18.8%) patients discontinued placebo in the neoadjuvant phase. In an exploratory mRNA-based subset analysis, patients with basal-like immune-activated tumors may have benefited from atezo. TNBC subtyping to identify basal-like immune-activated tumors and quantification of tumor-infiltrating lymphocytes to identify tumors with high tumor-infiltrating lymphocyte counts could be a promising strategy to identify patients who benefit from the addition of immune checkpoint inhibitors to neoadjuvant chemotherapy. ClinicalTrials.gov registration: NCT03281954 .
    DOI:  https://doi.org/10.1038/s41591-026-04565-6
  13. Oncoimmunology. 2026 Dec 31. 15(1): 2725500
      Sinonasal intestinal-type adenocarcinoma (ITAC) is a rare malignancy strongly associated with occupational exposure to wood dust that shares morphological and molecular similarities with colorectal adenocarcinoma. Standard treatments offer limited benefit for advanced or recurrent disease. It is therefore necessary to develop other strategies, such as immunotherapies, which require a better understanding of the tumor immune microenvironment, are needed. In this study, we investigated the immune microenvironment of ITACs (n=59), focusing on tumor-infiltrating T lymphocytes (TILs) and macrophages, in situ using immunohistochemistry, ex vivo by multiparametric flow cytometry, as well as by transcriptomic profiling. High CD8⁺ T-cell densities were associated with significantly improved disease-free and overall survival and emerged as an independent prognostic factor. These TILs express immune checkpoints (IC), the most highly expressed being PD1, TIGIT, and TIM-3. Furthermore, TIM-3 was not only expressed by TILs but also by immunoregulatory macrophages, and may represent an important immune escape mechanism in ITACs, since TIM-3 blockade in patient-derived explants led to increased tumor cell apoptosis. Tumor cells strongly expressed CD155 (TIGIT ligand) but not PDL1 (PD1 ligand); however expressed by myeloid cells. Finally, transcriptomic analyses identified one cluster displaying features of "hot tumors" characterized by adaptive immune activation, a better prognosis, and a higher immunotherapy response score, and another cluster with features of "cold tumors". These first exploratory and descriptive findings in ITACs provide a rationale for further investigations using ex vivo and in vivo models to evaluate the therapeutic potential of immunotherapies in these rare cancers.
    Keywords:  Immunotherapy; immune checkpoints; patient-derived explants; sinonasal intestinal type adenocarcinoma (ITAC); transcriptomics; tumor immune microenvironment
    DOI:  https://doi.org/10.1080/2162402X.2026.2725500
  14. Clin Transl Immunology. 2026 ;15(9): e70120
       Objectives: Despite their anti-cancer properties, natural killer (NK) cells are discarded during CAR T cell manufacturing. NK cells display promising responses as a cell therapy, exhibiting remarkable safety at high doses. Moreover, NK cells have complementary cytokine profiles and distinct antigen-recognition pathways to supplement CAR T cell therapies. We demonstrate parallel expansion of NK cells from the normally discarded PBMC of CAR T cell production for downstream combination therapies against solid and blood-borne malignancies.
    Methods: CAR T cells were manufactured from buffy coats with initial isolation of T cells from PBMC. NK cells were rapidly expanded in parallel by stimulating the remaining PBMC with membrane-bound (mb)IL-21 + mbIL-15 feeder cells. Expanded NK cells were assessed through spectral flow cytometry and applied as a pretreatment prior to CAR Tcell infusion against xenograft tumor models in NOD-scid IL2Rgammanull (NSG) mice.
    Results: Applying feeder cells to residual PBMC stimulated 5000-fold expansions of pure NK cells. Expanded NK cells expressed various activation receptors and displayed strong in vitro cytotoxicity, which was further improved in CAR T-conditioned medium. A pretreatment of NK cells enhanced existing CAR T cell therapies, improving in vivo tumor clearance and survival against breast cancer, acute lymphoblastic leukaemia and non-Hodgkin's lymphoma. While NK cells eliminated CD19-/- variants in vitro, they were unable to suppress antigen-escape relapse in vivo.
    Conclusion: This study encourages the optimal use of leukapheresis product. By recycling discarded PBMC into NK cells, CAR T cell therapies can be effectively enhanced, improving tumor clearance and survival.
    Keywords:  T cell; cell therapy; chimeric antigen receptor; manufacturing; natural killer
    DOI:  https://doi.org/10.1002/cti2.70120
  15. Front Immunol. 2026 ;17 1919543
      Tertiary lymphoid structures (TLS) have evolved from descriptive histological findings into candidate biomarkers and targetable immune niches in solid tumors. In intrahepatic cholangiocarcinoma (ICC), however, similar lymphoid architectures may have different biological implications depending on their relationship to tumor nests, immune circuitry, and suppressive stroma. We define a "TLS topotype" as the multidimensional state of an individual TLS-centered region, including the TLS core, its perilesional neighborhood, and the nearest tumor-stroma interface. This state integrates anatomic topology and tumor accessibility, structural maturity, putative effector circuitry, and suppressive context; treatment-induced plasticity is considered separately through longitudinal assessment. Because heterogeneous TLS may coexist within one tumor, the whole tumor is represented by a "topotype landscape" based on compartment-specific TLS density and the distribution of TLS-level states rather than a single label. Direct human ICC evidence supports compartment-resolved associations between TLS distribution and outcome and identifies suppressive B-cell, Treg, myeloid, and stromal programs. These observations establish spatial TLS organization as prognostically relevant, but do not validate the framework or demonstrate treatment-specific prediction. By contrast, TLS-resident TCF7-positive stem-like CD8 T-cell maintenance, coordinated B-cell-dendritic-cell-T-cell circuitry, and ICI-specific predictive value remain unproven in ICC and should be considered hypothesis-generating. The framework therefore distinguishes candidate circuit-competent, tumor-accessible TLS states from stromally excluded or suppressive-neighborhood states while allowing mixed profiles. Multiplex pathology and spatial transcriptomics may define these states, whereas radiomics, liquid biopsy, and pretreatment biopsy provide complementary but incomplete information. Biopsy can characterize only TLS states captured in sampled tissue, and radiomics currently estimates whole-lesion TLS probability rather than functional topology. Marker expression or spatial colocalization alone does not establish functional productivity, which requires evidence of tumor-antigen presentation or specificity, local lymphocyte renewal or clonal expansion, functional perturbation, or a spatially linked treatment response. By separating TLS-level state, tumor-level landscape, and patient-level trial stratum, this review provides a framework for biomarker development, therapeutic reprogramming, prospective validation, and longitudinal monitoring in ICC. No TLS topotype has yet been validated to predict differential benefit from immune checkpoint inhibition in ICC.
    Keywords:  immunotherapy; intrahepatic cholangiocarcinoma; radiomics; spatial immunology; stem-like CD8 T cells; tertiary lymphoid structures; translational medicine; tumor immune microenvironment
    DOI:  https://doi.org/10.3389/fimmu.2026.1919543
  16. Signal Transduct Target Ther. 2026 Sep 03. pii: 362. [Epub ahead of print]11(1):
      T cell exhaustion and T cell senescence constitute distinct yet partially overlapping differentiation states that collectively constrain T cell functionality. T cell exhaustion arises under conditions of chronic antigen exposure and is characterised by a progressive, hierarchical loss of effector capacity, sustained expression of inhibitory receptors, and extensive transcriptional, epigenetic and metabolic reprogramming. By contrast, T cell senescence represents a more stable and terminal state, driven by replicative history, age-associated decline or stress-induced damage, and is defined by durable cell cycle arrest, altered differentiation, metabolic remodelling and acquisition of a pro-inflammatory secretory phenotype. In the context of cancer, dysfunctional T cells contribute to tumour progression, while also representing a major barrier to the success of T cell-based immune therapies, which strongly rely on the fitness, persistence and functional plasticity of T cells. Although substantial efforts have focused on overcoming exhaustion and optimising T cell manufacturing, senescence remains comparatively underexplored and presents unique therapeutic challenges due to its relative resistance to functional reprogramming. This review provides a comprehensive overview of T cell replenishment in homeostasis, followed by the molecular hallmarks and signalling pathways of T cell senescence and exhaustion. We discuss the current landscape of T cell-based immune therapies, including immune checkpoint blockade, T cell engagers and adoptive cell therapies, and explain how T cell dysfunction impacts their therapeutic outcomes. Finally, we highlight emerging strategies to prevent or overcome T cell dysfunction in adoptive cell therapy products.
    DOI:  https://doi.org/10.1038/s41392-026-02923-x
  17. Res Sq. 2026 Aug 25. pii: rs.3.rs-10629841. [Epub ahead of print]
      Immune checkpoint inhibitors (ICIs) have improved outcomes for patients with melanoma and are now the standard of care for high-risk and advanced disease. However, long-term benefits are observed in only around 25% of patients, with significant risk for immune-related adverse events, highlighting the need for predictive biomarkers. To develop a minimally invasive, pre-treatment biomarker strategy, we profiled functional and subset-specific transcripts in peripheral blood T lymphocytes (PBTLs) and applied machine learning to identify predictive signatures. Patients were enrolled prior to receiving ICI monotherapy in the adjuvant (Exploratory n=61, Validation=78) or metastatic (Exploratory n=48, Validation=46) settings. Following feature selection, random forest models were trained and benchmarked against empirical null models. In the adjuvant setting, CD160 and GZMB predicted recurrence (95th percentile), while treatment-limiting toxicity was predicted by a signature comprising TNFRSF18, VTCN1, TIGIT, CCR4, and AHR (97th percentile). In the metastatic setting, baseline CD45RB, a marker of T-cell differentiation, most strongly predicted progression within one year (93.9th percentile). Distinct signatures in the adjuvant and metastatic settings suggest differences in T-cell programs associated with patient outcomes. These findings support further evaluation of pre-treatment circulating T-cell transcriptional profiles as predictors of ICI response and toxicity in melanoma.
    DOI:  https://doi.org/10.21203/rs.3.rs-10629841/v1
  18. Int J Biol Sci. 2026 ;22(13): 6985-7002
      Chimeric antigen receptor (CAR) T cell therapy has achieved remarkable clinical success in hematological malignancies. However, its efficacy in solid tumors such as lung cancer remains constrained by the immunosuppressive tumor microenvironment (TME). Aberrant vascular architecture and dense stroma constitute major physical barriers that hinder CAR T cell infiltration. Additionally, an immunosuppressive cellular network, dominated by myeloid-derived suppressor cells and tumor-associated macrophages, further restricts CAR T cell expansion and function. Moreover, immune checkpoint signaling, inhibitory cytokines, dysregulated chemokine gradients, and metabolic reprogramming under hypoxia collectively create a hostile biochemical and metabolic milieu that drives CAR T cell dysfunction and exhaustion. This review systematically outlines these multifactorial barriers within the lung cancer TME and discusses emerging strategies, including combinatorial approaches, engineered CAR T designs, and microenvironment-modulating platforms, that aim to improve the therapeutic efficacy of CAR T cell therapy in lung cancer.
    Keywords:  CAR T cell therapy; immunotherapy; lung cancer; solid tumors; tumor microenvironment
    DOI:  https://doi.org/10.7150/ijbs.131261
  19. Immune Netw. 2026 Aug;26(4): e29
      T cell-based universal influenza vaccines aim to generate robust lung-resident memory CD8+ T cells (TRM), yet it remains unclear whether conserved Ags elicit equally protective TRM pools. We compared recombinant adenoviral vectors expressing the influenza A nucleoprotein (rAd/NP) or polymerase basic protein 1 (PB1). After intranasal immunization, both induced comparable magnitudes of respiratory Ag-specific CD8+ T cells, yet only rAd/NP provided 100% survival after lethal influenza challenge. This disparity reflected qualitative differences in the TRM pool; nucleoprotein (NP)-specific cells were predominantly CD103-CD49a+, a phenotype associated with superior cytotoxicity, whereas PB1-specific cells were mainly CD103+CD49a-. Furthermore, NP-specific CD8+ T cells showed 100-fold higher functional avidity and stronger lung-local CTL activity than PB1-specific cells. Our findings demonstrate that TRM quality-specifically phenotypic bias and functional avidity-rather than magnitude, are the primary determinants of vaccine-mediated protection. This study underscores the critical importance of Ag selection in optimizing T cell-based universal vaccine strategies.
    Keywords:  Adenoviral vector; Nucleoprotein; Tissue resident memory T cells; Universal influenza vaccine
    DOI:  https://doi.org/10.4110/in.2026.26.e29